Combinatorial organic synthesis is becoming an important tool in drug discovery. Methods for the synthesis of large numbers of diverse compounds have been described (Ellman et al., Chem. Rev. 96, 555-600 (1996)), as have methods for tagging systems (Ohlmeyer et al., Proc. Natl. Acad. Sci. USA 90, 10922-10926 (1993)). The growing importance of combinatorial synthesis has created a need for new resins and linkers having chemical and physical properties that accommodate a wide range of conditions. Success in combinatorial synthesis on solid phase supports depends on the ability to synthesize diverse sets of molecules and to then cleave those molecules from the supports cleanly and in good yield.
Linkers are molecules that are attached to a solid support and to which the desired members of a library of chemical compounds may in turn be attached. When the construction of the library is complete, the linker allows clean separation of the target compounds from the solid support without harm to the compounds and preferably without damage to the support. Several linkers have been described in the literature. Their value is constrained by the need to have sufficient stability which allows the steps of combinatorial synthesis under conditions that will not cleave the linker. An additional constraint is the need to have a fairly high lability under at least one set of conditions that is not employed in the chemical synthesis.
For example, if an acid labile linker is employed, then the combinatorial synthesis must be restricted to reactions that do not require the presence of an acid of sufficient strength to endanger the integrity of the linker. Likewise, when a photocleavable linker is employed, conditions that exclude light are necessary to avoid untimely cleavage of the compound from the resin. This sort of balancing act often imposes serious constraints on the reactions that are chosen for preparation of the library.
The 4-[4-(hydroxymethyl)-3-methoxyphenoxy]butyryl residue is a known linker, which is attached to a solid support having amino groups by forming an amide with the carboxyl of the butyric acid chain. N-Protected amino acids are attached to the hydroxyl of the 4-hydroxymethyl group via their carboxyl to form 2,4-dialkoxybenzyl esters, which can be readily cleaved in acid media when the synthesis is complete (Riniker et al., Tetrahedron 49, 9307-9312 (1993)). The drawback to such 2,4-dialkoxybenzyl esters is their instability with many of the reagents that are available for use in combinatorial synthesis resulting in cleavage of the ester.
A somewhat more stable ester is formed from 4-[4-(hydroxymethyl)phenoxy]butyric acid, described in European published application EP 445915. In this case, the ester was cleaved with a 90:5:5 mixture of trifluoroacetic acid, dimethyl sulfide and thioanisole.
When the desired product is a peptide amide, the 4-[4-(formyl)-3,5-dimethoxyphenoxy]butyryl residue has been employed as a linker. This particular linker is attached to a solid phase substrate via the carboxyl of the butyric acid chain, and the 4-formyl group is reductively aminated. N-Protected amino acids are then reacted with the alkylamine via their carboxyl to form 2,4,6-trialkoxybenzylamides. These may be cleaved by 1:1 trifluoroacetic acid in dichloromethane (PCT application WO97/23508).
If a photocleavable linker is used to attach chemical compounds to the main support, milder photolytic conditions of cleavage can be used which complement traditional acidic or basic cleavage techniques. A wider range of combinatorial synthetic conditions will be tolerated by photocleavable linkers (Gallop et al., J. Med. Chem. 37, 1233-1251 (1994); Gordon et al., J. Med. Chem. 37, 1385-1401 (1994)).
A phenacyl based linking group that is photocleavable has been described (Wang et al., J. Org. Chem. 41, 3258 (1976)). The 4-bromomethyl-3-nitrobenzoyl residue has been widely employed as a photocleavable linker for both peptide acids and amides (Rich et al., J. Am. Chem. Soc. 97, 1575-1579 (1975); Hammer et al., Int. J. Peptide Protein Res. 36, 31-45 (1990)). This linker suffers from unduly slow cleavage rates, with typical photolysis times in organic solvents ranging from 12 to 24 hours. Moreover, photolytic cleavage of the linker generates a reactive and chromogenic nitrosoaldehyde on the resin support which can trap liberated compounds (Patchnornik et al., J. Am. Chein. Soc. 92, 6333-6335 (1970)). An .alpha.-methyl-2-nitrobenzyl linker was designed to obviate formation of the nitroso-aldehyde, but inefficient release of pentapeptides resulted due to swelling of the resin support (Ajajaghosh et al., Tetrahedron 44, 6661-6666 (1988)). Photocleavable linkers such as the 3-bromomethyl-4-nitro-6-methoxyphenoxyacetyl residue are stable to acidic or basic conditions yet, are rapidly cleavable under mild conditions and do not generate highly reactive byproducts (U.S. Pat. No. 5,739,386, issued Apr. 14, 1998).
It would be useful to have a linker-resin combination that would withstand a wider range of reaction conditions in combinatorial synthesis, but that could be readily and cleanly cleaved following completion of the solid phase synthesis.